Study On The Preparation Of Transgenic Quail By Liposome Transfection

Transgenic avian has a broad prospect, can provide humans with high yield and quality of meat and egg protein, provides biomedical and health care of protein and breeding for disease resistance. At present, preparation transgenic poultry of method has electric transfection method, reverse transcription virus and liposome transfection method, after analysis discussion and combined with the actual situation of our laboratory,we used liposome transfection blastoderm cells to confirm the feasibility of preparing transgenic quail, preliminary building transgenic quail of model, providing experimental basis and foundation for the best method for producing transgenic quails. The results are as follows:Translating the pEGFP-C2plasmid into the subgerminal cavity by liposome transfection blastoderm cells, the packing ratio of plasmid and liposome lipofection-2000optimal is1:2.5(μg:μL),the plasmid in liposome will not overflow by gel retardation of electrophoresis under this concentration. After formaldehyde fixation operation on the blastoderm, observed that injection of bromophenol blue in the subgerminal cavity.In this experiment,we injected the mixed liquid into the subgerminal cavity of quail egg blastoderm cells, verified by genomic PCR and detected pEGFP-C2plasmid found on the sixth day embryos. Confirmed that exogenous DNA can be injected into subgerminal cavity of stage X in quail egg and successful transfection blastoderm cells by this experimental injection method.verification of transgenic offspring showed gene expression in quail gizzard and proventriculus of visceral tissues in a newly hatched7day-old GO generation was detected by PCR, the results of sequencing and sequence alignment was correct; an adult GO transgenic female quails visceral tissues expressing green fluorescent protein; two GO generation transgenic quail appearance can be observed in the expression of green fluorescent protein; Three adult transgenic quail passed the transgene onto its offspring when mated to wild-type quail, The positive ratio was33.96%,20.59%, and10%, respectively. Then crossing in three adult transgenic quail, the Gi generation positive rate were10.87%and42.86%.In summary, transfection method in this experiment is usefull to translate exogenous DNA into the subgerminal cavity in quail, plasmid DNA after injection could be expressed in embryonic and adult offspring, and the exogenous gene can be integrated into the quail reproductive glands and their offspring.